Although it has been demonstrated that iNOS plays an important role in host defense against microbial pathogens, the exact function of iNOS in T cells and chronic inflammatory diseases has not been defined. TH17 cells, which secrete interleukin 17 (IL-17) and IL-22 comprise a recently identified subset of CD4+ T cells distinct from the TH1 and TH2 subsets and RORgt has been identified as a key transcription factor for TH17 cell differentiation. Increasing evidence indicates that TH17 immune responses are involved in the pathogenesis of various autoimmune/inflammatory diseases. Thus, blocking TH17 cell activation could lead to the development of novel strategies for the treatment of chronic inflammatory diseases. We show that iNOS knock out mice display more robust TH17 cell differentiation without major effects on either TH1 or TH2 cell lineages. We demonstrated that iNOS protein was induced in activated CD4+ T cells and the use of an iNOS selective inhibitor L-NIL significantly increased the percentage of IL-17-producing CD4+ T cells in WT cell cultures, while, an NO donor, SNAP, dose-dependently suppressed IL-17 production in WT and iNOS-/- T cell cultures. In addition, tyrosine residues of RORgt protein were nitrated resulting in the inhibition of RORgt-mediated IL-17 promoter activation. Finally, transfer of iNOS-/- CD4+CD45Rbhi cells into RAG-/- mice induces more severe colitis compared to control CD4+CD45Rbhi cells and mice reconstituted with iNOS-/- cells had a significantly higher percentage of IL-17-producing cells than control mice. The results suggest that T cell-derived iNOS negatively regulates the development of TH17 immune responses resulting in the control of inflammation. This proposal is structured around three aims: 1) We will characterize the molecular mechanisms involved in the regulation of TH17 cell differentiation by T cell-derived iNOS. 2) We will characterize the function of T cell- derived-iNOS on human TH17, TH1, TH2, and Treg cell development. 3) We will analyze the roles of iNOS in different cell compartments including macrophages, dendritic cells, and T cells in the development of colitis. The proposed studies will define a novel transcriptional inhibitor of TH17 cell differentiation and highlight th importance of T cell-derived iNOS as a novel therapeutic target for the treatment of chronic inflammatory diseases.